Article Figures & Data
Figures
- Figure 1.
Sex-specific differences in MU dynamics in SOD1G37R/YFP mice. A, Examples of MU arbors from a male and a female SOD1/YFP mice during three sessions (male: session 2, 3, and 5; female: session 2, 4, and 5), with higher magnification on-focus insets of NMJs of interest (digital zoom, dashed box in low magnification). Green: YFP-labeled axon; red: nAChR. Note how the imaged motor axon retracted from two NMJs (arrowheads) between sessions 3 and 5 in the male example. Furthermore, note the formation of a new axonal branch between sessions 2 and 4 in the female example (arrow), innervating an NMJ that previously did not belong to this MU (MU expansion). B, MU dynamic diagram of the MUs shown in A, showing that MU are much more likely to expand in female SOD1G37R/YFP mice. Black lines represent the axonal arborization and each box represents a single NMJ. Note how asynchronous NMJ losses occurred in both these MUs, but how partial losses seem more frequent in the male MU. C, D, Histograms showing the average proportion of NMJs from the initial pool which are innervated (green), re-innervated (purple), or lost (red) by the MU and the proportion which are gained (blue) for female (C) and male (D) SOD1G37R/YFP mice. Numbers in the histogram bars represent the number of remaining MU arbors (those which did not globally degenerate) over the total number of MUs observed. Again, note how MU expansions are more frequent in female than in male mice. Scale bar: 100 μm (low magnification) and 25 μm (high magnification).
- Figure 2.
Motor function declines slightly faster in female SOD1G37R/YFP mice. A, B, Body weight (A) and all-limb grip strength (B) curves of male and female (light and dark colors, respectively) WT/YFP and SOD1G37R/YFP mice (blue and red, respectively) during the symptomatic stages (WT female: N = 14; WT male: N = 6; SOD1 female: N = 8; SOD1 male: N = 12). Data are presented as mean ± SEM. Main effects and interactions are reported in Table 1 and in the text. Asterisks and crosses represent biologically relevant statistically significant differences in the post hoc test (Tukey’s multiple comparisons). Asterisks represent differences compared with the other genotype, while crosses represent differences compared with the other sex; */†p < 0.05, **/††p < 0.01, ***/†††p < 0.001, ****/††††p < 0.0001.
- Figure 3.
α-motor neuron loss and NMJ denervation are slightly more pronounced in female SOD1G37R/YFP mice at P360. A, Representative image of lumbar spinal cord sections from a female WT/YFP mice at P360 (red: ChAT; blue: NeuN; green: YFP). B, Representative image of NMJs in the tibialis anterior muscle of a female WT/YFP mice at P360 (red: α-BTX; green: NF-M and SV2). C, Representative examples of lumbar spinal cord sections from female (top) and male (bottom) SOD1G37R/YFP mice at P360. D, Representative images of NMJs in the tibialis anterior muscle of female (top) and male (bottom) SOD1G37R/YFP mice at P360. E, F, Quantification of the number of α-motor neurons (E, ChAT+ NeuN+ cells) and γ-motor neurons (F, ChAT+ NeuN– cells) per ventral horn at P360 (WT female: N = 4; WT male: N = 2; SOD1 female: N = 4; SOD1 male: N = 5). G, Quantification of the percentage of fully innervated NMJs in the tibialis anterior at P360 (WT female: N = 4, n = 1078; WT male: N = 2, n = 446; SOD1 female: N = 4, n = 977; SOD1 male: N = 5, n = 820). Data are presented as mean ± SEM. Main effects and interactions are reported in Table 2 and in the text. Asterisks represent biologically relevant statistically significant differences in the post hoc test (E, F, Tukey’s multiple comparisons; G, Holm–Sidak’s correction); *p < 0.05, **p < 0.01, ***p < 0.001. Scale bar: 100 μm.
Tables
Analysis Statistical test Main effect p value Body weight (Fig. 2A) RM three-way ANOVA Time <0.001*** Genotype <0.001*** Sex <0.001*** Time × genotype <0.001*** Time × sex 0.854 Sex × genotype 0.868 Time × sex × genotype 0.397 Grip strength (Fig. 2B) RM three-way ANOVA Time <0.001*** Genotype <0.001*** Sex <0.001*** Time × genotype <0.001*** Time × sex 0.007** Sex × genotype 0.023* Time × sex × genotype 0.011* Bold values represent statistically significant differences (p < 0.05).
- Table 2
Effect of sex and genotype on neuronal survival and NMJ innervation in SOD1G37R mice
Analysis Statistical Test Main effect p value α-Motor neurons counts
(Fig. 3E)Two-way ANOVA Genotype <0.001*** Sex 0.002** Sex × genotype 0.941 γ-Motor neurons counts
(Fig. 3F)Two-way ANOVA Genotype 0.673 Sex 0.042* Sex × genotype 0.365 NMJ innervation
(Fig. 3G)GLM, logistic distribution Genotype <0.001*** Sex 0.431 Sex × genotype 0.019* Bold values represent statistically significant differences (p < 0.05).
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